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M. Dolores Loureiro, M. Carmen Martínez, Jean-Michel Boursiquot, and Patrice This

`Albariño' (Vitis vinifera L.) is an important grape cultivar in Spain, morphologically diverse but subject to much misnaming. The objectives of the present work were to correct some of the more common misnamings concerning `Albariño' and to evaluate the genetic variability within this cultivar by analyzing DNA polymorphisms using randomly amplified polymorphic DNA (RAPD) markers and microsatellite techniques. Several accessions of `Albariño' (16 accessions from Misión Biológica de Galicia, one accession from El Encin, one accession from Rancho de la Merced), related cultivars (`Alvarinho', `Caíño blanco', `Cainho branco', `Loureiro'), and cultivars presumably identical to misnomers (`Savagnin blanc' and `Gewürztraminer') were analyzed using 20 RAPD markers and six microsatellite loci. Both techniques revealed polymorphism among `Albariño', `Caíño blanco', `Albariño' from Rancho de la Merced and `Loureiro'. No polymorphism was detected among the 16 `Albariño' accessions from Galicia, the `Albariño' accession from El Encin and `Alvarinho', nor among the `Albariño' accession from Rancho de la Merced, `Savagnin blanc' and `Gewürztraminer', nor between `Caíño blanco' and `Cainho branco'. These results enabled us to clarify the main misnomers concerning these cultivars. The absence of polymorphism among the true `Albariño' accessions did not allow the detection of any clonal variation. The suitability of both techniques for defining the cultivar level for grapevine is discussed.

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Soon O. Park, Dermot P. Coyne, and James R. Steadman

Bean rust, caused by Uromyces appendiculatus, is an important disease of common bean (Phaseolus vulgaris L.). The objective was to identify RAPD markers linked to the gene (Ur-6) for specific resistance to rust race 51 using bulked segregant analysis in an F2 segregating population from the common bean cross pinto `Olathe' (resistant to rust) × great northern Nebraska #1 selection 27 (susceptible to rust). A single dominant gene controlling specific resistance to race 51 was hypothesized based on F2 segregation, and then was confirmed in the F3 generation. A good fit to a 3:1 ratio for band presence to band absence for each of three markers was observed in 100 F2 plants. Three RAPD markers were detected in a coupling phase linkage with the Ur-6 gene. Coupling-phase RAPD marker OAB14.600 was the most closely linked to the Ur-6 gene at a distance of 3.5 cM among these markers. No RAPD markers were identified in a repulsion phase linkage with the Ur-6 gene. The RAPD markers linked to the gene for specific rust resistance of Middle American origin detected here, along with other independent rust resistance genes from other germplasm, could be utilized to pyramid multiple genes into a bean cultivar for more durable rust resistance.

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Martha Dávila, Dermot Coyne, Shree Singh, and Guenhwa Jung

The genes involved in F1 seedling abnormal development and lethality in inter-gene pool crosses have been designated as Dl1 (MesoAmerican=MA) and Dl2 (Andean=A) (Shii et al., 1980, J. Hered. 71:218–222). The different degrees of leaf crippling (C) in segregating populations of crosses was due to the interaction between the Dl1 or Dl2 loci, growing environment, and the lcr allele (Singh and Molina, 1996, J. Hered., In press). The objective was to identify RAPD markers linked to the genes for crippling (lcr) and seedling lethality (Dl) using the bulked segregation analysis procedure for F2 of MA × A crosses. Crosses were made between C lines, FB 10413-24-2, WA 7807-305, and TY 5578-220 and normal (N) parents and tester stocks for Dl1 and Dl2 genes. The F2 FB 10413-24-2 × Carioca segregated 13 N:3C. F3 families segregated 3N:1C. RAPD marker OPB-10 was linked to Lcr at 31.2 cM. F3 families segregated 1N:3C. RAPD marker OPO16 was linked to Dl1 at 27 cM. The F2 WA-7807-305 × Rio Tibagi segregated 3N:1C. RAPD marker OPS-03 was linked to Lcr at 32.6 cM.

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Hong Y. Yang, Schuyler S. Korban, Jutta Kruger, and Hanna Schmidt

Apple scab, caused by Venturia inaequalis (Cke.) Wint., is the most serious disease of apple trees. Resistance to V. inaequalis, derived from the small-fruited species Malus floribunda 821, is determined by a major dominant gene Vf. Our major objective is to identify RAPD markers linked to the Vf gene. The approach in this paper is based on the introgression of the Vf gene from M. floribunda into commercial cultivars. Almost 200 random sequence decamer-primers have been used to screen a pair of bulked samples and the donor parent M. floribunda clone 821 for markers linked to the Vf gene conferring resistance to apple scab. A single primer has been identified which generated a PCR fragment, OPK16/1300, from the donor parent M. floribunda clone 821 and the scab-resistant selections/cultivars bulk, but not from the scab-susceptible recurrent parent bulk. Co-segregation analysis using a segregating apple progeny and polymorphism analysis of individual scab-resistant Coop selections/cultivars have confirmed that this marker is linked to the scab-resistance gene Vf. OPK16/1300 has since been cloned and sequenced. Sequence-specific primers of 25 oligonucleotides based on the marker have been synthesized and used to screen further M. floribunda clone 821, scab-susceptible apple cultivars, scab-resistant apple cultivars, and scab-resistant Coop selections. The sequence-specific primers have identified polymorphisms of OPK16/1300 based on the presence or absence of a single band.

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Antonio Figueira, Jules Janick, and Peter Goldsbrough

RAPD markers were used to examine genetic similarity in cacao. DNA from 30 cacao cultivars amplified using 15 arbitrary oligonucleotide primers, produced a total of 112 fragments, of which 88% were polymorphic. A phenogram was developed which illustrates the genetic relationships among the cacao cultivars representing the four major geographic groups of cacao (Criollo, Trinitario, Forastero Lower Amazonian, and Forastero Upper Amazonian). The phenogram indicated a general separation of the four groups into three clusters. Criollos and Trinitarios (supposedly hybrids between Forastero and Criollos types) appeared in a single cluster. Lower Amazonian cultivars (mainly selections made in Bahia, Brazil) appeared in a separate cluster. The third cluster consisted of the Upper Amazonian cultivars, which were originally collected from the region believed to be the center of origin of this crop. This cluster displayed the furthest genetic distance from the others. Crosses between Upper Amazon germplasm and local selections have shown heterosis in clonal crosses, which has been exploited in all genetic improvement programs for cacao. We propose that genetic distances based on RAPD markers can be potentially used as a criterion to select parents capable of producing superior hybrids and populations. Genetic relationships can also be useful to define germplasm collections and conservation strategies. Studies are underway to compare phenograms derived from RAPD markers and ribosomal RNA gene polymorphisms.

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R.N. Trigiano, K.M. Kaveriappa, S.E. Schlarbaum, M.T. Windham, and W. Witte

DNA amplification fingerprinting (DAF) was Used to characterize both parents (different cultivars) in breeding experiments with Cornus florida. Putative hybrids were fingerprinted and true crosses identified by finding unique male parent products in amplification profiles. Both manual and honey bee mediated pollinations successfully produced hybrid seed. Axillary buds from seedlings were used to initiate proliferating shoot cultures on woody plant medium with 4.5 μm BA. Initiation and development of adventitious roots were dependent on IBA (4.1 μm), sucrose (0–2%), and agar (0.2–0.6%) concentrations. About 40–50% of the microshoots produced roots and were acclimatized to greenhouse conditions. Cultures have been maintained without loss of regeneration potential for over 2 years. Clonal material can be reentered into the breeding program or used to evaluate horticultural characteristics in different environments and locales.

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Yiqi Zhen, Zuozhou Li, Hongwen Huang, and Ying Wang

Forty-eight kiwifruit cultivars and selections, representing more than 90% of total world kiwifruit production, were investigated using nine SSR markers to establish genetic identities, and evaluate genetic diversity and relatedness. These nine SSRs were polymorphic and a total of 213 alleles were detected, resulting in a mean number of 23.7 alleles per locus, ranging from nine to 38 alleles. One hundred and thirty-three alleles were found to be common to both A. chinensis and A. deliciosa, while 33 and 36 were specific to A. chinensis and A. deliciosa, respectively. In addition, 34 alleles were specific to one single genotype and provided a set of valuable alleles for cultivar identification. A single SSR locus UDK 96-414 could differentiate all 48 genotypes except two presumable clones. Mean number of alleles per locus (A), percentage of polymorphic loci (P), and direct count heterozygosity (Ho) assessed for each genotype over all loci revealed considerable differences among these 48 genotypes. On average, A = 2.6, P = 89.4% and Ho = 0.546 were found in A. chinensis cultivars, while A = 3.5, P = 97.0% and Ho = 0.671 in A. deliciosa cultivars. Consensus fingerprint profiling using SSR markers is a useful and reliable method for establishing genetic identities of kiwifruit cultivars and selections. It also improves evaluation effectiveness of genetic diversity and relatedness compared to RAPD markers.

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Margaret R. Pooler, Louise G.H. Riedel, S.E. Bentz, and A.M. Townsend

Controlled pollinations were made between five hemlock (Tsuga) species from eastern North America and Asia, resulting in over 5700 germinating seedlings. A subset of putative hybrid seedlings from each cross was tested for authenticity by various DNA marker systems. The most reliable and useful system for verifying hybrids was amplified fragment-length polymorphism (AFLP) markers. Hybridizations between the eastern North American species, T. canadensis [L.] Carriere and T. caroliniana Engelm., and the Asian species, T. chinensis (Franch.) E. Pritz., were used as a model to test the inheritance, reliability, and ease of use of these markers. Using AFLP markers, we were able to verify 58 hybrids between T. caroliniana and T. chinensis, one hybrid between T. caroliniana and T. canadensis, but could find no definitive hybrids between T. canadensis and T. chinensis. Results using other marker systems, including RAPD, SCAR, ITS, and SSR, are also presented.

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W.V. Baird, R.E. Ballard, S. Rajapakse, and A.G. Abbott

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Kentaro Kitahara, Shogo Matsumoto, Toshiya Yamamoto, Junichi Soejima, Tetsuya Kimura, Hiromitsu Komatsu, and Kazuyuki Abe

We examined the genetic diversity and relatedness among apple (Malus ×domestica Borkh.) cultivars in Japan. The 42 apple cultivars, including major cultivars in Japan, were divided into five groups based on SSR genotypes. Most economically important cultivars belong in three groups: Fuji-Delicious, Golden Delicious, and Jonathan groups, and their genetic backgrounds seemed to be narrow. We also investigated the parent-offspring relationships of nine apple cultivars. `Jonathan', `Fuji', and `Rero 11' were identified as the respective paternal parents of three cultivars described as having unknown paternal parents (i.e., `Akagi', `Ambitious', and `Hokuto'). `Starking Delicious', `Senshu', and `Golden Delicious', rather than `Ralls Janet', `Hatsuaki', and `Indo', seemed to be the paternal parents of `Kinsei', `Kiou', and `Mellow', respectively. `Carolina Red June' was excluded as a paternal parent of `Ranzan'. Both attributed parents of `Scarlet' (`Akane' and `Starking Delicious') were excluded, and it was suggested that `Fuji' was used as either a maternal or a paternal parent of `Scarlet'. `Jonathan' rather than `McIntosh' seems to be a maternal parent of `Yukari'.